DETECTION OF ALPHA-THALASSEMIAS BY MULTIPLEX POLYMERASE CHAIN-REACTION

Although alpha-thalassemia is the most common genetic abnormality in t he world, there is currently no routine laboratory method to definitiv ely identify individuals who are affected. We describe a rapid and sim ple method that utilizes deletion-sensitive primers to amplify normal DNA sequences. Deletions involving the regions responsible for most of the alpha-thalassemia cases in the US prevent amplification with thes e primers. In tests with DNA isolated from small amounts (10 mu L) Of whole blood, the deletion-sensitive primers gave rise to the expected 248-and 375-bp (base pair) amplification products in normal individual s. These primers, along with primers designed to bind to a nonaffected control sequence from the hemoglobin beta chain, could be amplified s imultaneously (multiplex polymerase chain reaction). This made it poss ible to detect heterozygotes for alpha-thalassemia-2 (one alpha locus deleted) by determining the ratios of the 248- and 375-bp amplificatio n products to the product of the control sequence (268 bp). The method is rapid and simple and can be performed in a routine clinical labora tory.